US6316214B1 - ETM-775 metabolite of ecteinascidin 743 - Google Patents

ETM-775 metabolite of ecteinascidin 743 Download PDF

Info

Publication number
US6316214B1
US6316214B1 US09/309,947 US30994799A US6316214B1 US 6316214 B1 US6316214 B1 US 6316214B1 US 30994799 A US30994799 A US 30994799A US 6316214 B1 US6316214 B1 US 6316214B1
Authority
US
United States
Prior art keywords
etm
compounds
spectrum
sioh
ecteinascidin
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Fee Related
Application number
US09/309,947
Inventor
Kenneth L. Rinehart
Jose J. Morales
Joel Reid
Isabel Reymundo
Pablo Floriano
Lola Garcia Gravalos
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Pharmamar SA
Original Assignee
University of Illinois
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by University of Illinois filed Critical University of Illinois
Priority to US09/309,947 priority Critical patent/US6316214B1/en
Assigned to BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS, THE reassignment BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS, THE ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: MORALES, JOSE J., RINEHART, KENNETH L.
Priority to US09/971,852 priority patent/US6867334B2/en
Application granted granted Critical
Publication of US6316214B1 publication Critical patent/US6316214B1/en
Assigned to PHARMA MAR, S.A. reassignment PHARMA MAR, S.A. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS, THE
Priority to US11/009,237 priority patent/US7115743B2/en
Anticipated expiration legal-status Critical
Expired - Fee Related legal-status Critical Current

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D515/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen, oxygen, and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00
    • C07D515/22—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen, oxygen, and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00 in which the condensed system contains four or more hetero rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D217/00—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems
    • C07D217/02—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems with only hydrogen atoms or radicals containing only carbon and hydrogen atoms, directly attached to carbon atoms of the nitrogen-containing ring; Alkylene-bis-isoquinolines
    • C07D217/10—Quaternary compounds
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D317/00—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms
    • C07D317/08—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3
    • C07D317/44—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D317/46—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems condensed with one six-membered ring
    • C07D317/48—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring
    • C07D317/62—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to atoms of the carbocyclic ring
    • C07D317/64—Oxygen atoms

Definitions

  • Et ecteinascidins
  • Et's are exceedingly potent antitumor agents isolated from the marine tunicate Ecteinascidia turbinata.
  • Et's 729, 743 and 722 have demonstrated promising efficacy in vivo, including activity against P388 murine leukemia, B16 melanoma, Lewis lung carcinoma, and several human tumor xenograft models in mice.
  • Ecteinascidin 743 (Et 743) has the following structure:
  • ETM ETM
  • ETM 305 and ETM 775 were isolated from a metabolic mixture obtained from a biochemical study performed by the Analytical Chemistry Department at PharmaMar, Spain. A similar metabolic study carried out by the Mayo Clinic led to the identification of ETM 204.
  • the structures of these ecteinascidin metabolites are as follows:
  • FIG. 1 is the 1 H NMR spectrum (500 MHz) of ETM-SiOH-1 (non-polar impurity) in CDCl 3 ;
  • FIG. 2 is the HPLC chromatogram of ETM-SiOH-4 (ETM 775);
  • FIG. 3 is the HPLC chromatogram of ETM-SiOH-3 (ETM 305);
  • FIG. 4 is the HPLC chromatogram of ETM-SiOH-2 (trace metabolites).
  • FIG. 5 is the LRFAB mass spectrum of ETM 305 in M.B. (magic bullet 4 );
  • FIG. 6 is the ESI mass spectrum of ETM 305
  • FIG. 7 is the 1 H NMR spectrum (750 MHz) of ETM 305 in CD 3 OD;
  • FIG. 8 is the FAB/MS/MS spectrum of ETM 305.
  • FIG. 9 is the UV spectrum of ETM 305.
  • FIG. 10 is the UV spectrum of ETM
  • FIG. 11 is the LRFAB mass spectrum of ETM 775 in M.B.
  • FIG. 12 is the ESI mass spectrum of ETM 775 (positive mode).
  • FIG. 13 is the ESI mass spectrum of ETM 775 (negative mode).
  • FIG. 14 is the FAB/MS/MS spectrum of ETM 775 (m/z 138-302);
  • FIG. 15 is the FAB/MS/MS spectrum of ETM 775 (m/z 440-620);
  • FIG. 16 is the 1 H NMR spectrum (750 MHz) of ETM 775 in CD 3 OD;
  • FIG. 17 is the UV spectrum of ETM 775
  • FIG. 18 is the HPLC choromatogram of ETM 305.
  • FIG. 19 is the UV spectrum of ETM 305
  • FIG. 20 is the ESI mass spectrum of ETM 305.
  • FIG. 21 is the ESI mass spectrum of ETM 204
  • FIG. 22 is the 1 H NMR spectrum (500 MHz) of ETM 204 in CD 3 OD.
  • FIG. 23 is the ESI/MS/MS spectrum of ETM 204.
  • Et-743 (50 ⁇ M) was incubated with 0.4 mg/ml of human lymphoblast-expressed CYP3A4 isoform (Gentest Corporation, Woburn, Mass.) in 0.1 M Tris-HCl buffer (pH 7.4) containing an NADPH generating system (0.4 mM NADP + , 25 mM glucose-6-phosphate, 0.5 U/ml glucose-6-phosphate dehydrogenase and 3.3 mM magnesium chloride). After four (4) hours at 37° C., the reaction was stopped with ice cold acetonitrile and the solids removed by centrifugation (12,000 g, 4 min.). Supernatants were analyzed by HPLC.
  • ETM-SiOH-1 a lipid not structurally related to Et 743 as revealed by the 1 H NMR spectrum (FIG. 1 ).
  • cytotoxic fractions were further purified by HPLC (Phenomenex-Ultracarb ODS, 10 ⁇ m, 10 ⁇ 150 mm, 3:1 MeOH/H 2 O 0.02 M NaCl, 1 mL/min., Da Detection: 210, 220, 254 and 280 nm).
  • HPLC HPLC
  • ETM-SiOH-4, 0.2 mg yield 0. 1 mg of ETM 775 (FIG. 2 ).
  • ETM-SiOH-3 yield 0.3 mg of ETM 305 (FIG. 3 )
  • ETM-SiOH-2 consisted of a complex mixture of trace metabolites (FIG. 4 ).
  • ETM-SiOH fractions R f , weight and cytoxic activity.
  • L1210 growth Test inhibition tube (%) at ID # # R f a Weight 500 ng/mL ETM-SiOH 1 1 0.9 2.0 mg 0 ETM-SiOH 2 2 0.5, 0.7 0.3 mg 80 b ETM-SiOH 3 4-5 0.5 0.4 mg 30 ETM-SiOH 4 6 0.3 0.2 mg 3 a Silica gel TLC using 9:1 CHCl 3 /MeOH as mobile phase. b 30% inhibition at 250 ng.
  • ETM 305 (IC 50 0.2 ⁇ m/mL vs L1210 cells) showed a molecular ion at 306.0977 by HRFAB/MS (FIG. 5 ). This data is in agreement with the molecular formula C 15 H 16 NO 6 ( ⁇ 0.1 mmu). ESI/MS analysis confirmed the molecular weight of ETM 305 (FIG. 6 ); a molecular ion at m/z 306 was observed together with its sodium adduct (m/z 328). The 1 H NMR spectrum of ETM 305 (FIG. 7) was very important for the structural assignment. Resonances at ⁇ 2.04, 2.28 and 6.09 were almost identical to those of Me-6 ( ⁇ 2.03), —OCOCH 3 ( ⁇ 2.29) and the dioxy-methylene protons ( ⁇ 6.11 and 6.01) in Et 743, 1 respectively.
  • ETM 775 (IC 50 0.2 ⁇ g/mL vs L1210 cells) showed a molecular ion at 776.2489 by HRPAB/MS FIG. 11 ). This data is in agreement with the molecular formula C 39 H 42 N 3 O 12 S ( ⁇ 0.0 mmu) which indicated that ETM 775 is an oxidation product of Et 743. Both, positive and negative mode ESI/MS spectra confirmed the molecular weight of ETM 775 (FIGS. 12 and 13 ). Because of the limited amount of ETM 775, the structural assignment was carried out mainly by interpretation of its mass spectral data.
  • the ETM sample was filtered through a C18 sep-pack and the eluant (3:1 MeOH/H 2 O) concentrated under a nitrogen stream. Purification of the resulting residue by HPLC (same conditions as described above) revealed the presence of a compound with a retention time identical to that of ETM 305 (FIG. 18 ). Both, the UV (FIG. 19) and ESI/MS (FIG. 20) spectra of M1 were identical to that of ETM 305. Thus, it was concluded that M1 metabolite had the same chemical structure as ETM 305.
  • the provided sample was filtered through a C18 sep-pack and the eluant (3:1 MeOH/H 2 O) concentrated under a nitrogen stream and the resulting residue analyzed by FAB/MS, ESI/MS and 1 H NMR.
  • ETM 204 showed a molecular ion at 204.1024 by HRFAB/MS. This data is in agreement with the molecular formula C 12 H 14 NO 2 ( ⁇ 0.0 mmu). ESI/MS analysis confirmed the molecular weight as 204 (FIG. 21 ). The molecular formula matched with the molecular formula of the a-unit in Et 743. Thus, the chemical structure of ETM 204 was proposed to be the aromatic ammonium salt derivative shown in Scheme 3. This simple compound (as well as the other metabolites) can easily be monitored to assay the breakdown of Et 743 in vivo.
  • the ESI/MS/MS of ETM 204 (FIG. 23) showed a prominent peak ion at 189 corresponding to the apparent loss of the N-methyl group (204—CH 3 ).
  • the present invention includes bioactive compounds, and accordingly, an embodiment of the present invention is directed to methods of treatment using such compounds.
  • the compounds of the present invention have exhibited in vitro cytoxicity against tumor cell lines. It is anticipated that these in vitro activities will likewise extend to in vivo utility.
  • compositions containing the active compounds identified herein and methods of treatment employment such pharmaceutical compositions are provided.
  • the active compounds of the present invention exhibit antitumor activity, thus, the present invention also provides a method of treating any mammal affected by a malignant tumor sensitive to these compounds, which comprises administering to the affected individual a therapeutically effective amount of an active compound or mixture of compounds, or pharmaceutical compositions thereof.
  • the present invention also relates to pharmaceutical preparations, which contain as active ingredient one or more of the compounds of this invention, as well as the processes for its preparation.
  • Example of pharmaceutical compositions include any solid (tablets, pills, capsules, granules, etc.) or liquid (solutions, suspensions of emulsions) with suitable composition or oral, topical or parenteral administration, and they may contained the pure compound or in combination with any carrier of other pharmacologically active compounds. These compositions may need to be sterile when administered parenterally.
  • unit dose refers to physically discrete units suitable as unitary dosages for animals, each unit containing a predetermined quantity of active material calculated to produce the desired antitumor effect in association with the required diluent; i.e., carrier, or vehicle.
  • the specifications for the novel unit dose of this invention are dictated by and are directly dependent on (a) the unique characteristics of the active material and the particular antitumor effect to be achieved, and (b) the limitations inherent in the art of compounding such active material for antitumor use in animals.
  • Unit dosage forms are typically prepared from the frozen or dried active compound (or salts thereof) by dispersement in a physiologically tolerable (i.e., acceptable) diluent or vehicle such as water, saline or phosphate-buffered saline to form an aqueous composition.
  • a physiologically tolerable (i.e., acceptable) diluent or vehicle such as water, saline or phosphate-buffered saline to form an aqueous composition.
  • diluents are well known in the art and are discussed, for example, in Remington's Pharmaceutical Sciences, 16th Ed., Mack Publishing Company, Easton, Pa. (1980) at pages 1465-1467.
  • Dosage forms can also include an adjuvant as part of the diluent.
  • adjuvants such as complete Freund's adjuvant (CFA), incomplete Freund's adjuvant (IFA) and alum are materials well known in the art, and are available commercially from several sources.
  • the quantity of active compound to be administered depends, inter alia, on the animal species to be treated, the subject animal's size, the size of the tumor (if known), the type of tumor (e.g., solid) present, and the capacity of the subject to utilize the active compound. Precise amounts of active compound required to be administered depend on the judgment of the practitioner and are peculiar to each individual, particularly where humans are the treated animals. Dosage ranges, however, can be characterized by a therapeutically effective blood concentration and can range from a concentration of from about 0.01 ⁇ M to about 100 ⁇ M, preferably about 0.1 ⁇ M to 10 ⁇ M.
  • Suitable regimes for initial administration and booster injections are also variable, but are typified by an initial administration followed by repeated doses at one or more hour intervals by a subsequent injection or other administration.
  • continuous intravenous infusion sufficient to maintain a therapeutically effective concentration in the blood are contemplated.

Landscapes

  • Organic Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oncology (AREA)
  • Hematology (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Nitrogen Condensed Heterocyclic Rings (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Heterocyclic Compounds That Contain Two Or More Ring Oxygen Atoms (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The purification and structure elucidation of several products of the metabolism of Et 743 by human cytochrome CYP3A4 have been accomplished. These compounds are abbreviated herein as “ETM” followed by a numeric value, which represents the approximate molecular weight. Three compounds have been identified to date, namely ETM 305, ETM 775 and ETM 204. The structures of these ecteinascidin metabolites are as follows:
Figure US06316214-20011113-C00001

Description

CROSS-REFERENCE TO RELATED APPLICATION
This application claims priority benefit from copending U.S. Provisional Application Ser. No. 60/085,024, filed May 11, 1998, the disclosure of which is hereby incorporated herein by reference.
BACKGROUND OF THE INVENTION
The ecteinascidins (herein abbreviated Et or Et's) are exceedingly potent antitumor agents isolated from the marine tunicate Ecteinascidia turbinata. In particular, Et's 729, 743 and 722 have demonstrated promising efficacy in vivo, including activity against P388 murine leukemia, B16 melanoma, Lewis lung carcinoma, and several human tumor xenograft models in mice.
The isolation and characterization of natural Et 743 is taught in U.S. Pat. No. 5,089,273 which is hereby incorporated herein by reference. The preparation of synthetic Et 743 is taught in U.S. Pat. No. 5,721,362, which is hereby incorporated herein by reference.
The antitumor activities of ecteinascidin compounds, particularly Et 729 and Et 743 are well documented in the scientific literature. See for example, Goldwasser et al., Proceedings of the American Association for Cancer Research, 39: 598 (1998); Kuffel et al., Proceedings of the American Association for Cancer Research, 38: 596 (1997); Moore et al., Proceedings of the American Association for Cancer Research, 38: 314 (1997); Mirsalis et al., Proceedings of the American Association for Cancer Research, 38: 309 (1997); Reid et al., Cancer Chemotherapy and Pharmacology, 38: 329-334 (1996); Faircloth et al., European Journal of Cancer, 32A, Supp. 1, pp. S5 (1996); Garcia-Rocha et al., British Journal of Cancer, 73: 875-883 (1996); Eckhardt et al., Proceedings of the American Association for Cancer Research, 37: 409 (1996); Hendriks et al., Proceedings of the American Association for Cancer Research, 37: 389 (1996); the disclosures of which are hereby incorporated herein by reference.
Ecteinascidin 743 (Et 743) has the following structure:
Figure US06316214-20011113-C00002
In view of the impressive antitumor activities of this class of compounds, the search continues for related structures that may possess equal or higher levels of antitumor activity. The present invention, which is directed to the isolation and characterization of natural metabolites of Et 743, is a result of these continued studies.
SUMMARY OF THE INVENTION
The purification and structure elucidation of several products of the metabolism of Et 743 by human cytochrome CYP3A4 have been accomplished. These compounds are abbreviated herein as “ETM” followed by a numeric value which represents the approximate molecular weight.
For example, ETM 305 and ETM 775 were isolated from a metabolic mixture obtained from a biochemical study performed by the Analytical Chemistry Department at PharmaMar, Spain. A similar metabolic study carried out by the Mayo Clinic led to the identification of ETM 204. The structures of these ecteinascidin metabolites are as follows:
Figure US06316214-20011113-C00003
BRIEF DESCRIPTION OF THE DRAWINGS
The present invention may be better understood by reference to the drawings accompanying this specification, wherein:
FIG. 1 is the 1H NMR spectrum (500 MHz) of ETM-SiOH-1 (non-polar impurity) in CDCl3;
FIG. 2 is the HPLC chromatogram of ETM-SiOH-4 (ETM 775);
FIG. 3 is the HPLC chromatogram of ETM-SiOH-3 (ETM 305);
FIG. 4 is the HPLC chromatogram of ETM-SiOH-2 (trace metabolites);
FIG. 5 is the LRFAB mass spectrum of ETM 305 in M.B. (magic bullet4);
FIG. 6 is the ESI mass spectrum of ETM 305;
FIG. 7 is the 1H NMR spectrum (750 MHz) of ETM 305 in CD3OD;
FIG. 8 is the FAB/MS/MS spectrum of ETM 305;
FIG. 9 is the UV spectrum of ETM 305;
FIG. 10 is the UV spectrum of ETM;
FIG. 11 is the LRFAB mass spectrum of ETM 775 in M.B.;
FIG. 12 is the ESI mass spectrum of ETM 775 (positive mode);
FIG. 13 is the ESI mass spectrum of ETM 775 (negative mode);
FIG. 14 is the FAB/MS/MS spectrum of ETM 775 (m/z 138-302);
FIG. 15 is the FAB/MS/MS spectrum of ETM 775 (m/z 440-620);
FIG. 16 is the 1H NMR spectrum (750 MHz) of ETM 775 in CD3OD;
FIG. 17 is the UV spectrum of ETM 775;
FIG. 18 is the HPLC choromatogram of ETM 305;
FIG. 19 is the UV spectrum of ETM 305;
FIG. 20 is the ESI mass spectrum of ETM 305;
FIG. 21 is the ESI mass spectrum of ETM 204;
FIG. 22 is the 1H NMR spectrum (500 MHz) of ETM 204 in CD3OD; and
FIG. 23 is the ESI/MS/MS spectrum of ETM 204.
DETAILED DESCRIPTION OF THE INVENTION
I. Et 743 Metabolic Study
A. Preparation of Metabolic Mixture—ETM
Et-743 (50 μM) was incubated with 0.4 mg/ml of human lymphoblast-expressed CYP3A4 isoform (Gentest Corporation, Woburn, Mass.) in 0.1 M Tris-HCl buffer (pH 7.4) containing an NADPH generating system (0.4 mM NADP+, 25 mM glucose-6-phosphate, 0.5 U/ml glucose-6-phosphate dehydrogenase and 3.3 mM magnesium chloride). After four (4) hours at 37° C., the reaction was stopped with ice cold acetonitrile and the solids removed by centrifugation (12,000 g, 4 min.). Supernatants were analyzed by HPLC.
B. Purification of ETM 305 and ETM 775
2.6 mg of ETM (generated as in A, above) was dissolved in a small amount of CHCl3 and loaded into a silica gel column (8×100 mm glass column filled with a silica gel/CHCl3 slurry). First, the column was eluted with CHCl3 followed by CHCl3/MeOH mixtures (98, 96, 94, 92 and 90%). A total of ten test tubes were collected (3 mL each) and combined on the basis of TLC to yield four fractions (Table 1). The less polar and non-cytotoxic fraction (ETM-SiOH-1, 2 mg) consisted of a lipid not structurally related to Et 743 as revealed by the 1H NMR spectrum (FIG. 1).
The remaining cytotoxic fractions were further purified by HPLC (Phenomenex-Ultracarb ODS, 10 μm, 10×150 mm, 3:1 MeOH/H2O 0.02 M NaCl, 1 mL/min., Da Detection: 210, 220, 254 and 280 nm). The most polar fraction (ETM-SiOH-4, 0.2 mg) yield 0. 1 mg of ETM 775 (FIG. 2). ETM-SiOH-3 yield 0.3 mg of ETM 305 (FIG. 3), and ETM-SiOH-2 consisted of a complex mixture of trace metabolites (FIG. 4).
TABLE 1
ETM-SiOH fractions: Rf, weight and cytoxic activity.
L1210 growth
Test inhibition
tube (%) at
ID # # Rf a Weight 500 ng/mL
ETM-SiOH 1 1 0.9 2.0 mg  0
ETM-SiOH 2 2 0.5, 0.7 0.3 mg 80b
ETM-SiOH 3 4-5 0.5 0.4 mg 30
ETM-SiOH 4 6 0.3 0.2 mg  3
aSilica gel TLC using 9:1 CHCl3/MeOH as mobile phase.
b30% inhibition at 250 ng.
C. The Structure of ETM 305
ETM 305 (IC50 0.2 μm/mL vs L1210 cells) showed a molecular ion at 306.0977 by HRFAB/MS (FIG. 5). This data is in agreement with the molecular formula C15H16NO6(Δ0.1 mmu). ESI/MS analysis confirmed the molecular weight of ETM 305 (FIG. 6); a molecular ion at m/z 306 was observed together with its sodium adduct (m/z 328). The 1H NMR spectrum of ETM 305 (FIG. 7) was very important for the structural assignment. Resonances at δ2.04, 2.28 and 6.09 were almost identical to those of Me-6 (δ2.03), —OCOCH3 (δ2.29) and the dioxy-methylene protons (δ6.11 and 6.01) in Et 743,1 respectively.
In addition, it was observed resonances corresponding to a —CH═CH—NHCHO unit (δ7.09, d, 1H, J=15 Hz; δ6.19, d, 1H, J=15 Hz; δ8.04, s, 1H),2 and an additional methyl group (δ2.52, s, 3H). The chemical shift of this methyl group match pretty well wit that of the methyl group on acetophenone3 (δ2.55). It is interesting to note that the 1H NMR spectrum of ETM 305 consisted of two sets of resonances (4:1 ratio) due to rotational conformers around the —NH—CHO bond The 1H NMR data together with the MS data suggested that ETM 305 had the B-unit aromatic ring system of Et 743 attached to a vinyl-formamide unit and to a methyl ketone as shown in Scheme 1. FAB/MS/MS on m/z 306 supported the proposed structure (FIG. 8).
Figure US06316214-20011113-C00004
D. The Structure of ETM 775
ETM 775 (IC50 0.2 μg/mL vs L1210 cells) showed a molecular ion at 776.2489 by HRPAB/MS FIG. 11). This data is in agreement with the molecular formula C39H42N3O12S (Δ0.0 mmu) which indicated that ETM 775 is an oxidation product of Et 743. Both, positive and negative mode ESI/MS spectra confirmed the molecular weight of ETM 775 (FIGS. 12 and 13). Because of the limited amount of ETM 775, the structural assignment was carried out mainly by interpretation of its mass spectral data. FABMS/MS on M+H of ETM 775 (m/z 776) was critical in assigning the location of the extra oxygen was located on N-2 in the form of an N-oxide as revealed by peaks at m/z 276 and 260 (276-oxygen). A fragment ion at m/z 232, not observed in Et 743, suggested that the carbinol amine oxygen was oxidized to the amide (Scheme 3). The structures of the A- and C-units in ETM 775 remained intact as revealed by the presence of the characteristic mass spectral peaks at m/z 204 (A-unit), and m/z 224 and 250 (C-unit).1 Both, the 750 750 Mhz 1H NMR (FIG. 16) and the UV (FIG. 17) spectra resembled those of Et 743.1
Figure US06316214-20011113-C00005
II. Et 743—Mayo Metabolic Study
A. M1 metabolite (ETM 305)
The ETM sample was filtered through a C18 sep-pack and the eluant (3:1 MeOH/H2O) concentrated under a nitrogen stream. Purification of the resulting residue by HPLC (same conditions as described above) revealed the presence of a compound with a retention time identical to that of ETM 305 (FIG. 18). Both, the UV (FIG. 19) and ESI/MS (FIG. 20) spectra of M1 were identical to that of ETM 305. Thus, it was concluded that M1 metabolite had the same chemical structure as ETM 305.
B. M2 metabolite (ETM 204)
The provided sample was filtered through a C18 sep-pack and the eluant (3:1 MeOH/H2O) concentrated under a nitrogen stream and the resulting residue analyzed by FAB/MS, ESI/MS and 1H NMR.
C. The Structure of ETM 204 (M2)
ETM 204 showed a molecular ion at 204.1024 by HRFAB/MS. This data is in agreement with the molecular formula C12H14NO2 (Δ0.0 mmu). ESI/MS analysis confirmed the molecular weight as 204 (FIG. 21). The molecular formula matched with the molecular formula of the a-unit in Et 743. Thus, the chemical structure of ETM 204 was proposed to be the aromatic ammonium salt derivative shown in Scheme 3. This simple compound (as well as the other metabolites) can easily be monitored to assay the breakdown of Et 743 in vivo.
Figure US06316214-20011113-C00006
A 1H NMR spectrum (FIG. 22) of ETM 204 showed resonances that supported the proposed structure: four aromatics signals (δ9.2, s; δ7.8, d, J=5 Hz, and 6 6.8, s) and three methyl singlets (δ4.2, δ3.9 and δ2.4) The ESI/MS/MS of ETM 204 (FIG. 23) showed a prominent peak ion at 189 corresponding to the apparent loss of the N-methyl group (204—CH3).
Biological Studies of ETM-305 and ETM-775
Compounds ETM-305 and ETM-775 have been assayed employing standard protocols for the following tumor cell lines; P-388 (murine leukemia); A-549 (human lung carcinoma); HT-29 (human colon adenocarcinoma); and MEL-28 (human malignant melanoma). See, for example, Bergeron et al., Biochem. Biophys. Res. Comm., 1984, 121 (3) 848-854 and Schroeder et al., J. Med. Chem., 1981, 24 1078-1083. These results are shown below in Table 2:
TABLE 2
Cell Line & Activity
IC50 (μg/ml)
Compound:
P-388 A-549 HT-29 MEL-28
ETM-305 0.5  0.5  0.5  0.25
ETM-775 0.01 0.01 0.01 0.01
Methods of Treatment
The present invention includes bioactive compounds, and accordingly, an embodiment of the present invention is directed to methods of treatment using such compounds. As described above, the compounds of the present invention have exhibited in vitro cytoxicity against tumor cell lines. It is anticipated that these in vitro activities will likewise extend to in vivo utility.
These compounds have been isolated in substantially pure form, i.e., at a purity level sufficient to allow physical and biological characterization thereof. These compounds have been found to possess specific antitumor activities and as such they will be useful as medicinal agents in mammals, particularly in humans. thus, another aspect of the present invention concerns pharmaceutical compositions containing the active compounds identified herein and methods of treatment employment such pharmaceutical compositions.
As described above, the active compounds of the present invention exhibit antitumor activity, thus, the present invention also provides a method of treating any mammal affected by a malignant tumor sensitive to these compounds, which comprises administering to the affected individual a therapeutically effective amount of an active compound or mixture of compounds, or pharmaceutical compositions thereof. The present invention also relates to pharmaceutical preparations, which contain as active ingredient one or more of the compounds of this invention, as well as the processes for its preparation.
Example of pharmaceutical compositions include any solid (tablets, pills, capsules, granules, etc.) or liquid (solutions, suspensions of emulsions) with suitable composition or oral, topical or parenteral administration, and they may contained the pure compound or in combination with any carrier of other pharmacologically active compounds. These compositions may need to be sterile when administered parenterally.
The terms “unit dose” as it pertains to the present invention refers to physically discrete units suitable as unitary dosages for animals, each unit containing a predetermined quantity of active material calculated to produce the desired antitumor effect in association with the required diluent; i.e., carrier, or vehicle. The specifications for the novel unit dose of this invention are dictated by and are directly dependent on (a) the unique characteristics of the active material and the particular antitumor effect to be achieved, and (b) the limitations inherent in the art of compounding such active material for antitumor use in animals.
Unit dosage forms are typically prepared from the frozen or dried active compound (or salts thereof) by dispersement in a physiologically tolerable (i.e., acceptable) diluent or vehicle such as water, saline or phosphate-buffered saline to form an aqueous composition. Such diluents are well known in the art and are discussed, for example, in Remington's Pharmaceutical Sciences, 16th Ed., Mack Publishing Company, Easton, Pa. (1980) at pages 1465-1467.
Dosage forms can also include an adjuvant as part of the diluent. Adjuvants such as complete Freund's adjuvant (CFA), incomplete Freund's adjuvant (IFA) and alum are materials well known in the art, and are available commercially from several sources.
The quantity of active compound to be administered depends, inter alia, on the animal species to be treated, the subject animal's size, the size of the tumor (if known), the type of tumor (e.g., solid) present, and the capacity of the subject to utilize the active compound. Precise amounts of active compound required to be administered depend on the judgment of the practitioner and are peculiar to each individual, particularly where humans are the treated animals. Dosage ranges, however, can be characterized by a therapeutically effective blood concentration and can range from a concentration of from about 0.01 μM to about 100 μM, preferably about 0.1 μM to 10 μM.
Suitable regimes for initial administration and booster injections are also variable, but are typified by an initial administration followed by repeated doses at one or more hour intervals by a subsequent injection or other administration. Alternatively, continuous intravenous infusion sufficient to maintain a therapeutically effective concentration in the blood are contemplated.
REFERENCES
The following background references are provided to assist the reader in understanding this invention. To the extent necessary, the contents are hereby incorporated herein by reference.
1. A) Rinehart et al., J. Org. Chem. 1990, 55, 4512. B) Rinehart et al., J. Am. Chem. Soc., 1996, 118 9017.
2. Herbert et al., J. Chem. Soc. Perkin Trans. I, 1987, 1593.
3. Pretsch et al. Tables of Spectral Data for Structure Determination of Organic Compounds; Springer-Verla: Berlin, 1989; p. H125.
4. Rinehart et al., Biochem. Res. Commun., 1984, 124, 350.
The present invention has been described in detail, including the preferred embodiments thereof However, it will be appreciated that those skilled in the art, upon consideration of the present disclosure, may make modifications and/or improvements on this invention and still be within the scope and spirit of this invention.

Claims (2)

What is claimed is:
1. Substantially pure ETM-775 having the following structure:
Figure US06316214-20011113-C00007
2. A pharmaceutical composition comprising ETM-775 and a pharmaceutically acceptable carrier, diluent or excipient.
US09/309,947 1998-05-11 1999-05-11 ETM-775 metabolite of ecteinascidin 743 Expired - Fee Related US6316214B1 (en)

Priority Applications (3)

Application Number Priority Date Filing Date Title
US09/309,947 US6316214B1 (en) 1998-05-11 1999-05-11 ETM-775 metabolite of ecteinascidin 743
US09/971,852 US6867334B2 (en) 1998-05-11 2001-10-03 Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US11/009,237 US7115743B2 (en) 1998-05-11 2004-12-10 Metabolites of ecteinascidin 743

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US8502498P 1998-05-11 1998-05-11
US09/309,947 US6316214B1 (en) 1998-05-11 1999-05-11 ETM-775 metabolite of ecteinascidin 743

Related Child Applications (1)

Application Number Title Priority Date Filing Date
US09/971,852 Continuation US6867334B2 (en) 1998-05-11 2001-10-03 Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4

Publications (1)

Publication Number Publication Date
US6316214B1 true US6316214B1 (en) 2001-11-13

Family

ID=22188976

Family Applications (3)

Application Number Title Priority Date Filing Date
US09/309,947 Expired - Fee Related US6316214B1 (en) 1998-05-11 1999-05-11 ETM-775 metabolite of ecteinascidin 743
US09/971,852 Expired - Fee Related US6867334B2 (en) 1998-05-11 2001-10-03 Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US11/009,237 Expired - Fee Related US7115743B2 (en) 1998-05-11 2004-12-10 Metabolites of ecteinascidin 743

Family Applications After (2)

Application Number Title Priority Date Filing Date
US09/971,852 Expired - Fee Related US6867334B2 (en) 1998-05-11 2001-10-03 Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US11/009,237 Expired - Fee Related US7115743B2 (en) 1998-05-11 2004-12-10 Metabolites of ecteinascidin 743

Country Status (22)

Country Link
US (3) US6316214B1 (en)
EP (1) EP1077698B1 (en)
JP (2) JP4583598B2 (en)
KR (1) KR100643092B1 (en)
CN (3) CN100339375C (en)
AT (1) ATE289810T1 (en)
AU (1) AU759281B2 (en)
BG (1) BG65235B1 (en)
BR (1) BR9910419A (en)
CA (1) CA2331593C (en)
CZ (1) CZ302168B6 (en)
DE (1) DE69923965T2 (en)
ES (1) ES2239442T3 (en)
HU (1) HUP0101960A3 (en)
IL (2) IL139607A0 (en)
NO (1) NO318081B1 (en)
NZ (1) NZ508585A (en)
PL (1) PL192409B1 (en)
RU (1) RU2225864C2 (en)
SK (1) SK285069B6 (en)
TR (1) TR200003470T2 (en)
WO (1) WO1999058125A1 (en)

Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20020032326A1 (en) * 1998-05-11 2002-03-14 Rinehart Kenneth L. Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US20030216397A1 (en) * 2000-04-12 2003-11-20 Maria Flores Antitumoral ecteinascidin derivatives
US20040002602A1 (en) * 1999-05-14 2004-01-01 Andres Francesch Synthetic process for the manufacture of an ecteinaschidin compound
US20040019056A1 (en) * 2000-05-15 2004-01-29 Ignacio Manzanares Anititumoral analogs of et-743
US20040204419A1 (en) * 2000-11-03 2004-10-14 Myers Andrew G. Saframycins, analogues and uses thereof
US20060106021A1 (en) * 2001-07-17 2006-05-18 Valentin Martinez New antitumoral derivatives of et-743
US20060111570A1 (en) * 2000-05-15 2006-05-25 Pharma Mar, S.A., A Madrid, Spain Corporation Synthetic process for the manufacture of an ecteinaschidin compound
US20060128711A1 (en) * 2001-08-07 2006-06-15 Pilar Gallego Antitumoral analogs
US20070093658A1 (en) * 2002-02-04 2007-04-26 Pharma Mar, S.A. Synthesis of naturally occcuring ecteinascidins and related compounds
US20080146580A1 (en) * 2000-04-12 2008-06-19 Pharmamar Anititumoral Ecteinascidin Derivatives
US20080273822A1 (en) * 2003-03-21 2008-11-06 Lynn Bich-Quy Le Method And System For Withstanding A Shock Event For A Fluid Dynamic Bearing Motor
US10538535B2 (en) 2017-04-27 2020-01-21 Pharma Mar, S.A. Antitumoral compounds
US11590129B2 (en) 2010-11-12 2023-02-28 Pharma Mar, S.A. Combination therapy with an antitumor alkaloid

Families Citing this family (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
MY164077A (en) * 1999-05-13 2017-11-30 Pharma Mar Sa Compositions and uses of et743 for treating cancer
NZ525730A (en) * 2000-11-06 2004-12-24 Pharma Mar S Compositions for antitumour treatments containing Ecteinascidin 743
PT1689404E (en) 2003-11-13 2008-12-15 Pharma Mar Sau Combination of et-743 with 5-fluorouracil pro-drugs for the treatment of cancer
PL1658848T3 (en) 2004-10-29 2007-12-31 Pharma Mar Sa Formulations comprising ecteinascidin and a disaccharide
US20090325971A1 (en) * 2005-03-02 2009-12-31 The Trustees Of Columbia University In The City Of New York Pentacyclic Alkaloid Compounds and Methods of Use Thereof
GB0522082D0 (en) 2005-10-31 2005-12-07 Pharma Mar Sa Formulations
US20090012157A1 (en) 2006-02-06 2009-01-08 Sears Barry D Sesamol Derivatives as Novel Inhibitors of Arachidonic Acid Formation

Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4273773A (en) 1979-09-24 1981-06-16 American Home Products Corporation Antihypertensive tricyclic isoindole derivatives
US5089273A (en) 1986-06-09 1992-02-18 Board Of Trustees Of The University Of Illinois Ecteinascidins 729, 743, 745, 759A, 759B and 770
US5256663A (en) * 1986-06-09 1993-10-26 The Board Of Trustees Of The University Of Illinois Compositions comprising ecteinascidins and a method of treating herpes simplex virus infections therewith
US5459141A (en) 1992-06-19 1995-10-17 Hoechst Aktiengesellschaft Compounds 31668P and 31668U, a process for their production and their use
US5484717A (en) 1993-11-10 1996-01-16 American Cyanamid Company Antibiotic 31F508α1, 31F508α2, 31F508β1, 31F508β2
US5654426A (en) * 1993-12-02 1997-08-05 Univ Illinois Ecteinascidins
US5721362A (en) * 1996-09-18 1998-02-24 President And Fellows Of Harvard College Process for producing ecteinascidin compounds
WO1998046080A1 (en) 1997-04-15 1998-10-22 The Board Of Trustees Of The University Of Illinois Nucleophile substituted ecteinascidins and n-oxide ecteinascidins

Family Cites Families (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS59225189A (en) 1983-06-03 1984-12-18 Shionogi & Co Ltd Quinonamin derivative and its preparation
JPS6084288A (en) 1983-10-13 1985-05-13 Shionogi & Co Ltd Cyanoquinonamine acetate and its production
ATE69234T1 (en) 1986-06-09 1991-11-15 Univ Illinois ECTEINASCIDINE 729, 743, 745, 759A, 759B AND 770.
US5149804A (en) 1990-11-30 1992-09-22 The Board Of Trustees Of The University Of Illinois Ecteinascidins 736 and 722
US5809563A (en) * 1996-11-12 1998-09-15 Institute For The Development Of Emerging Architectures, Llc Method and apparatus utilizing a region based page table walk bit
CA2327468C (en) * 1998-04-06 2008-05-06 The Board Of Trustees Of The University Of Illinois Semi-synthetic ecteinascidins
US6316214B1 (en) * 1998-05-11 2001-11-13 The Board Of Trustees Of The University Of Illinois ETM-775 metabolite of ecteinascidin 743
US6124292A (en) 1998-09-30 2000-09-26 President And Fellows Of Harvard College Synthetic analogs of ecteinascidin-743
MY130271A (en) 1999-05-14 2007-06-29 Pharma Mar Sa Hemisynthetic method and new compounds
WO2001053299A1 (en) 2000-01-19 2001-07-26 The Trustees Of Columbia University In The City Of New York Compounds of the saframycin-ecteinascidin series, uses, and synthesis thereof
PT1280809E (en) 2000-04-12 2005-11-30 Pharma Mar Sa ECTEINASCIDINE ANTITUMER DERIVATIVES
MXPA02011319A (en) 2000-05-15 2003-06-06 Pharma Mar Sa Antitumoral analogs of et 743.

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4273773A (en) 1979-09-24 1981-06-16 American Home Products Corporation Antihypertensive tricyclic isoindole derivatives
US5089273A (en) 1986-06-09 1992-02-18 Board Of Trustees Of The University Of Illinois Ecteinascidins 729, 743, 745, 759A, 759B and 770
US5256663A (en) * 1986-06-09 1993-10-26 The Board Of Trustees Of The University Of Illinois Compositions comprising ecteinascidins and a method of treating herpes simplex virus infections therewith
US5459141A (en) 1992-06-19 1995-10-17 Hoechst Aktiengesellschaft Compounds 31668P and 31668U, a process for their production and their use
US5484717A (en) 1993-11-10 1996-01-16 American Cyanamid Company Antibiotic 31F508α1, 31F508α2, 31F508β1, 31F508β2
US5654426A (en) * 1993-12-02 1997-08-05 Univ Illinois Ecteinascidins
US5721362A (en) * 1996-09-18 1998-02-24 President And Fellows Of Harvard College Process for producing ecteinascidin compounds
WO1998046080A1 (en) 1997-04-15 1998-10-22 The Board Of Trustees Of The University Of Illinois Nucleophile substituted ecteinascidins and n-oxide ecteinascidins
US5985876A (en) * 1997-04-15 1999-11-16 Univ Illinois Nucleophile substituted ecteinascidins and N-oxide ecteinascidins

Cited By (41)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7115743B2 (en) * 1998-05-11 2006-10-03 Pharma Mar, S.A. Metabolites of ecteinascidin 743
US6867334B2 (en) * 1998-05-11 2005-03-15 Pharma Mar, S.A. Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US20050261326A1 (en) * 1998-05-11 2005-11-24 Pharma Mar, S.A., An Illinois Corporation Metabolites of ecteinascidin 743
US20020032326A1 (en) * 1998-05-11 2002-03-14 Rinehart Kenneth L. Metabolites of ecteinascidin 743 formed by human cytochrome CYP3A4
US20040002602A1 (en) * 1999-05-14 2004-01-01 Andres Francesch Synthetic process for the manufacture of an ecteinaschidin compound
US7524956B2 (en) 1999-05-14 2009-04-28 Pharma Mar, S.A. Hemisynthetic method and new compounds
US20080045713A1 (en) * 1999-05-14 2008-02-21 Pharma Mar, S.A. Hemisynthetic Method and New Compounds
US7241892B1 (en) 1999-05-14 2007-07-10 Pharma Mar, S.A. Hemisynthetic method and new compounds
US20030216397A1 (en) * 2000-04-12 2003-11-20 Maria Flores Antitumoral ecteinascidin derivatives
US7919493B2 (en) 2000-04-12 2011-04-05 Pharma Mar, S.A. Anititumoral ecteinascidin derivatives
US20080146580A1 (en) * 2000-04-12 2008-06-19 Pharmamar Anititumoral Ecteinascidin Derivatives
US7202361B2 (en) 2000-04-12 2007-04-10 Pharmamar Antitumoral ecteinascidin derivatives
US20060111570A1 (en) * 2000-05-15 2006-05-25 Pharma Mar, S.A., A Madrid, Spain Corporation Synthetic process for the manufacture of an ecteinaschidin compound
US7410969B2 (en) 2000-05-15 2008-08-12 Pharma Mar, S.A. Antitumoral analogs of ET-743
US8012975B2 (en) 2000-05-15 2011-09-06 Pharma Mar, S.A. Antitumoral analogs of ET-743
US20040019056A1 (en) * 2000-05-15 2004-01-29 Ignacio Manzanares Anititumoral analogs of et-743
US7420051B2 (en) 2000-05-15 2008-09-02 Pharma Mar, S.A. Synthetic process for the manufacture of an ecteinaschidin compound
US7247629B2 (en) 2000-05-15 2007-07-24 Pharma Mar, S.A. Antitumoral analogs of et-743
US20060019960A1 (en) * 2000-05-15 2006-01-26 Pharma Mar, S.A. Antitumoral analogs of ET-743
US20080051407A1 (en) * 2000-05-15 2008-02-28 Pharma Mar, S.A., A Madrid Spain Corporation Antitumoral analogs of ET-743
US6809099B2 (en) 2000-11-03 2004-10-26 President And Fellows Of Harvard College Saframycins, analogues and uses thereof
US20040204419A1 (en) * 2000-11-03 2004-10-14 Myers Andrew G. Saframycins, analogues and uses thereof
US7122549B2 (en) 2000-11-03 2006-10-17 President And Fellows Of Harvard College Saframycins, analogues and uses thereof
US20100197695A1 (en) * 2001-07-17 2010-08-05 Valentin Martinez Antitumoral derivatives of et-743
US20060106021A1 (en) * 2001-07-17 2006-05-18 Valentin Martinez New antitumoral derivatives of et-743
US8076337B2 (en) 2001-07-17 2011-12-13 Pharma Mar, S.A. Antitumoral derivatives of ET-743
US7759345B2 (en) 2001-07-17 2010-07-20 Pharma Mar, S.A. Antitumoral derivatives of ET-743
US20060128711A1 (en) * 2001-08-07 2006-06-15 Pilar Gallego Antitumoral analogs
US7763615B2 (en) 2001-08-07 2010-07-27 Pharma Mar, S.A. Ecteinascidin analogs for use as antitumour agents
US7795260B2 (en) 2002-02-04 2010-09-14 Pharma Mar, S.A. Synthesis of naturally occurring ecteinascidins and related compounds
US20100216987A1 (en) * 2002-02-04 2010-08-26 Valentin Martinez Barrasa Synthesis of naturally occuring ecteinascidins and related compounds
US7767659B2 (en) 2002-02-04 2010-08-03 Pharma Mar, S.A. Synthesis of naturally occurring ecteinascidins and related compounds
US20070093658A1 (en) * 2002-02-04 2007-04-26 Pharma Mar, S.A. Synthesis of naturally occcuring ecteinascidins and related compounds
US7947671B2 (en) 2002-02-04 2011-05-24 Pharma Mar, S.A. Synthesis of naturally occuring ecteinascidins and related compounds
US20090163507A1 (en) * 2002-02-04 2009-06-25 Pharma Mar, S.A. Synthesis of naturally occuring ecteinascidins and related compounds
US20080273822A1 (en) * 2003-03-21 2008-11-06 Lynn Bich-Quy Le Method And System For Withstanding A Shock Event For A Fluid Dynamic Bearing Motor
US11590129B2 (en) 2010-11-12 2023-02-28 Pharma Mar, S.A. Combination therapy with an antitumor alkaloid
US10538535B2 (en) 2017-04-27 2020-01-21 Pharma Mar, S.A. Antitumoral compounds
US11332480B2 (en) 2017-04-27 2022-05-17 Pharma Mar, S.A. Antitumoral compounds
US11339180B2 (en) 2017-04-27 2022-05-24 Pharma Mar, S.A. Antitumoral compounds
US11713325B2 (en) 2017-04-27 2023-08-01 Pharma Mar, S.A. Antitumoral compounds

Also Published As

Publication number Publication date
EP1077698B1 (en) 2005-03-02
US20050261326A1 (en) 2005-11-24
AU4073799A (en) 1999-11-29
SK285069B6 (en) 2006-05-04
CN1195514C (en) 2005-04-06
US6867334B2 (en) 2005-03-15
US20020032326A1 (en) 2002-03-14
US7115743B2 (en) 2006-10-03
AU759281B2 (en) 2003-04-10
JP2010215656A (en) 2010-09-30
ES2239442T3 (en) 2005-09-16
WO1999058125A1 (en) 1999-11-18
NO20005671D0 (en) 2000-11-10
DE69923965T2 (en) 2006-04-06
CN100548988C (en) 2009-10-14
ATE289810T1 (en) 2005-03-15
NZ508585A (en) 2002-11-26
TR200003470T2 (en) 2001-06-21
NO20005671L (en) 2001-01-10
CZ20004183A3 (en) 2001-06-13
RU2225864C2 (en) 2004-03-20
EP1077698A4 (en) 2002-05-08
EP1077698A1 (en) 2001-02-28
BR9910419A (en) 2001-01-09
DE69923965D1 (en) 2005-04-07
KR20010071237A (en) 2001-07-28
HUP0101960A2 (en) 2001-11-28
NO318081B1 (en) 2005-01-31
CZ302168B6 (en) 2010-11-24
JP4583598B2 (en) 2010-11-17
PL192409B1 (en) 2006-10-31
BG65235B1 (en) 2007-09-28
KR100643092B1 (en) 2006-11-10
CA2331593C (en) 2008-08-05
HUP0101960A3 (en) 2003-03-28
PL344183A1 (en) 2001-10-08
CA2331593A1 (en) 1999-11-18
CN1371279A (en) 2002-09-25
IL139607A (en) 2006-12-10
CN1896064A (en) 2007-01-17
IL139607A0 (en) 2002-02-10
BG105023A (en) 2001-08-31
SK17152000A3 (en) 2001-07-10
JP2002514597A (en) 2002-05-21
CN100339375C (en) 2007-09-26
CN1683299A (en) 2005-10-19

Similar Documents

Publication Publication Date Title
US6316214B1 (en) ETM-775 metabolite of ecteinascidin 743
US5985876A (en) Nucleophile substituted ecteinascidins and N-oxide ecteinascidins
US5654426A (en) Ecteinascidins
EP0605622B1 (en) Oxygen substituted derivatives of nucleophile-nitric oxide adducts as nitric oxide donor prodrugs
EP1067933B1 (en) Semi-synthetic ecteinascidins
US20090247533A1 (en) Ecteinascidins
US4268676A (en) Mitomycin analogs
EP0507938A1 (en) Bis-(hydroxyalkylamino)-anthraquinone inhibitors of protein kinase c
US5360818A (en) 1,3-dioxane derivatives having protein kinase C inhibitory activity
MXPA00011064A (en) Metabolites of ecteinascidin 743
KR100533244B1 (en) Cytotoxic Alkaloid Derivatives Including Asmarine A and B Isolated from a Sponge
MXPA99009503A (en) Nucleophile substituted ecteinascidins and n-oxide ecteinascidins
WO1994010161A1 (en) 1,3-dioxane derivatives having protein kinase c inhibitory activity

Legal Events

Date Code Title Description
AS Assignment

Owner name: BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS, T

Free format text: ASSIGNMENT OF ASSIGNORS INTEREST;ASSIGNORS:RINEHART, KENNETH L.;MORALES, JOSE J.;REEL/FRAME:011381/0728

Effective date: 20001115

AS Assignment

Owner name: PHARMA MAR, S.A., SPAIN

Free format text: ASSIGNMENT OF ASSIGNORS INTEREST;ASSIGNOR:BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS, THE;REEL/FRAME:012652/0910

Effective date: 20011214

FPAY Fee payment

Year of fee payment: 4

FPAY Fee payment

Year of fee payment: 8

REMI Maintenance fee reminder mailed
LAPS Lapse for failure to pay maintenance fees
STCH Information on status: patent discontinuation

Free format text: PATENT EXPIRED DUE TO NONPAYMENT OF MAINTENANCE FEES UNDER 37 CFR 1.362

FP Lapsed due to failure to pay maintenance fee

Effective date: 20131113